Aryl and Heteroaryl Propene Amides
Cancer remains a leading cause of mortality in the United States and in the world. To be useful, a new chemotherapeutic agent should have a wide spectrum of activity and significant therapeutic index.
Aryl and heteroaryl propene amides have been prepared by reacting substituted aromatic acryloyl halides (such as cinnamoyl chlorides) with amino-substituted aromatic compounds (Kim et al., J. Chem. Soc. Perkin Trans. 2, 1995, p2257 describes reaction of cinnamoyl chlorides with anilines Alberghina et al., J. Org. Chem., Vol. 43, No. 6, 1978, p1122 describes reaction of furylacryloyl and thienylacryloyl chlorides with anilines.). Another route to aryl and heteroaryl propene amides involves reaction of an aromatic vinylhalide (preferably a bromide or iodide) with an aromatic amine using palladium catalysis (U.S. Pat. No. 3,988,358).
Certain propene amides have shown activity in treatment of atherosclerosis (Japanese patent 2001139550). Certain propene amides have also shown biological activity as inhibitors of oviposition in several herbivorous insect species (Blaakmeer et al, Journal of Natural Products, Vol. 57, No. 1, pg. 90, 1994, and Vol. 57, No. 8, pg. 1145). Some substituted cinnamanilides have demonstrated bacteriostatic properties (U.S. Pat. No. 3,975,435). Certain ortho-acyl -substituted cinnamanilides have been shown to inhibit the proliferation of vascular intimal cells which occurs incident to vascular stenosis (EP 0 855 387 A1). Other ortho acyl cinnamanilides have demonstrated biological activity as antiallergy agents (U.S. Pat. No. 4,337,270). N-imidazolyl cinnamides bearing an ortho-acyl substituent have been investigated as neovascularization inhibitors (Japanese patent applications 09363359 and 10307760). Other related compounds, N-indolylcinnamides having an ortho acyl substituent, have been shown to possess activity as COX-2 inhibitors (U.S. Pat. No. 6,300,363 B1). At least one cinnamanilide has been shown to be an antagonist at the platelet-activating factor (PAF) receptor (Lamotte-Brasseur et al., Lipids, Vol. 26, No. 12, 1991, p1167). Also N-pyridyl cinnamides have demonstrated antagonist activity of the enzyme CADH (Duran et al., Bull. Soc. Chim. Fr., 1987, No. 4, 1987, p 672).
Cell antiproliferative agents, and anticancer therapeutics in particular, are needed which are useful in inhibiting proliferation of and/or killing cancer cells. In particular, such agents are needed which are selective in the killing of proliferating cells such as tumor cells, but not normal cells. Antineoplastic agents are needed which are effective against a broad range of tumor types.
Ionizing Radiation Health Risks
Ionizing radiation has an adverse effect on cells and tissues, primarily through cytotoxic effects. In humans, exposure to ionizing radiation occurs primarily through therapeutic techniques (such as anticancer radiotherapy) or through occupational and environmental exposure.
A major source of exposure to ionizing radiation is the administration of therapeutic radiation in the treatment of cancer or other proliferative disorders. Depending on the course of treatment prescribed by the treating physician, multiple doses may be received by a subject over the course of several weeks to several months.
Therapeutic radiation is generally applied to a defined area of the subject's body which contains abnormal proliferative tissue, in order to maximize the dose absorbed by the abnormal tissue and minimize the dose absorbed by the nearby normal tissue. However, it is difficult (if not impossible) to selectively administer therapeutic ionizing radiation to the abnormal tissue. Thus, normal tissue proximate to the abnormal tissue is also exposed to potentially damaging doses of ionizing radiation throughout the course of treatment.
There are also some treatments that require exposure of the subject's entire body to the radiation, in a procedure called “total body irradiation”, or “TBI.” The efficacy of radiotherapeutic techniques in destroying abnormal proliferative cells is therefore balanced by associated cytotoxic effects on nearby normal cells. Because of this, radiotherapy techniques have an inherently narrow therapeutic index which results in the inadequate treatment of most tumors. Even the best radiotherapeutic techniques may result in incomplete tumor reduction, tumor recurrence, increasing tumor burden, and induction of radiation resistant tumors.
Numerous methods have been designed to reduce normal tissue damage while still delivering effective therapeutic doses of ionizing radiation. These techniques include brachytherapy, fractionated and hyperfractionated dosing, complicated dose scheduling and delivery systems, and high voltage therapy with a linear accelerator. However, such techniques only attempt to strike a balance between the therapeutic and undesirable effects of the radiation, and full efficacy has not been achieved.
For example, one treatment for subjects with metastatic tumors involves harvesting their hematopoietic stem cells and then treating the subject with high doses of ionizing radiation. This treatment is designed to destroy the subject's tumor cells, but has the side effect of also destroying their normal hematopoietic cells. Thus, a portion of the subject's bone marrow (containing the hematopoietic stem cells), is removed prior to radiation therapy. Once the subject has been treated, the autologous hematopoietic stem cells are returned to their body.
However, if tumor cells have metastasized away from the tumor's primary site, there is a high probability that some tumor cells will contaminate the harvested hematopoietic cell population. The harvested hematopoietic cell population may also contain neoplastic cells if the subject suffers from cancers of the bone marrow such as the various French-American-British (FAB) subtypes of acute myelogenous leukemias (AML), chronic myeloid leukemia (CML), or acute lymphocytic leukemia (ALL). Thus, the metastasized tumor cells or resident neoplastic cells must be removed or killed prior to reintroducing the stem cells to the subject. If any living tumorigenic or neoplastic cells are reintroduced into the subject, they can lead to a relapse.
Prior art methods of removing tumorigenic or neoplastic cells from harvested bone marrow are based on a whole-population tumor cell separation or killing strategy, which typically does not kill or remove all of the contaminating malignant cells. Such methods include leukopheresis of mobilized peripheral blood cells, immunoaffinity-based selection or killing of tumor cells, or the use of cytotoxic or photosensitizing agents to selectively kill tumor cells. In the best case, the malignant cell burden may still be at 1 to 10 tumor cells for every 100,000 cells present in the initial harvest (Lazarus et al. J. of Hematotherapy, 2(4):457-66, 1993).
Thus, there is needed a purging method designed to selectively destroy the malignant cells present in the bone marrow, while preserving the normal hematopoietic stem cells needed for hematopoietic reconstitution in the transplantation subject.
Exposure to ionizing radiation can also occur in the occupational setting. Occupational doses of ionizing radiation may be received by persons whose job involves exposure (or potential exposure) to radiation, for example in the nuclear power and nuclear weapons industries. Military personnel stationed on vessels powered by nuclear reactors, or soldiers required to operate in areas contaminated by radioactive fallout, risk similar exposure to ionizing radiation. Occupational exposure may also occur in rescue and emergency personnel called in to deal with catastrophic events involving a nuclear reactor or radioactive material. Other sources of occupational exposure may be from machine parts, plastics, and solvents left over from the manufacture of radioactive medical products, smoke alarms, emergency signs, and other consumer goods. Occupational exposure may also occur in persons who serve on nuclear powered vessels, particularly those who tend the nuclear reactors, in military personnel operating in areas contaminated by nuclear weapons fallout, and in emergency personnel who deal with nuclear accidents. Environmental exposure to ionizing radiation may also result from nuclear weapons detonations (either experimental or during wartime), discharges of actinides from nuclear waste storage and processing and reprocessing of nuclear fuel, and from naturally occurring radioactive materials such as radon gas or uranium. There is also increasing concern that the use of ordnance containing depleted uranium results in low-level radioactive contamination of combat areas.
Radiation exposure from any source can be classified as acute (a single large exposure) or chronic (a series of small low-level, or continuous low-level exposures spread over time). Radiation sickness generally results from an acute exposure of a sufficient dose, and presents with a characteristic set of symptoms that appear in an orderly fashion, including hair loss, weakness, vomiting, diarrhea, skin burns and bleeding from the gastrointestinal tract and mucous membranes. Genetic defects, sterility and cancers particularly bone marrow cancer) often develop over time. Chronic exposure is usually associated with delayed medical problems such as cancer and premature aging. An acute a total body exposure of 125,000 millirem may cause radiation sickness. Localized doses such as are used in radiotherapy may not cause radiation sickness, but may result in the damage or death of exposed normal cells.
For example, an acute total body radiation dose of 100,000-125,000 millirem (equivalent to 1 Gy) received in less than one week would result in observable physiologic effects such as skin burns or rashes, mucosal and GI bleeding, nausea, diarrhea and/or excessive fatigue. Longer term cytotoxic and genetic effects such as hematopoietic and immunocompetent cell destruction, hair loss (alopecia), gastrointestinal, and oral mucosal sloughing, venoocclusive disease of the liver and chronic vascular hyperplasia of cerebral vessels, cataracts, pneumonites, skin changes, and an increased incidence of cancer may also manifest over time. Acute doses of less than 10,000 millirem (equivalent to 0.1 Gy) typically will not result in immediately observable biologic or physiologic effects, although long term cytotoxic or genetic effects may occur.
A sufficiently large acute dose of ionizing radiation, for example 500,000 to over 1 million millirem (equivalent to 5-10 Gy), may kill a subject immediately. Doses in the hundreds of thousands of millirems may kill within 7 to 21 days from a condition called “acute radiation poisoning.” Reportedly, some of the Chernobyl firefighters died of acute radiation poisoning, having received acute doses in the range of 200,000-600,000 millirem (equivalent to 2-6 Gy). Acute doses below approximately 200,000 millirem do not result in death, but the exposed subject will likely suffer long-term cytotoxic or genetic effects as discussed above.
Acute occupational exposures usually occur in nuclear power plant workers exposed to accidental releases of radiation, or in fire and rescue personnel who respond to catastrophic events involving nuclear reactors or other sources of radioactive material. Suggested limits for acute occupational exposures in emergency situations were developed by the Brookhaven National Laboratories, and are given in Table 1.
TABLE 1Whole BodyConditions forDose LimitActivity RequiredConditions for Exposure 10,000 millirem*Protect propertyVoluntary, when lowerdose not practical 25,000 milliremLifesaving Operation;Voluntary, when lowerProtect General Publicdose not practical>25,000 milliremLifesaving operation;Voluntary, when lowerProtect largedose not practical, andpopulationthe risk has been clearlyexplained*100,000 millirem equals one sievert (Sv). For penetrating radiation such as gamma radiation, one Sv equals approximately one Gray (Gy). Thus, the dosage in Gy can be estimated as 1 Gy for every 100,000 millirem.
A chronic dose is a low level (i.e., 100-5000 millirem) incremental or continuous radiation dose received over time. Examples of chronic doses include a whole body dose of ˜5000 millirem per year, which is the dose typically received by an adult working at a nuclear power plant. By contrast, the Atomic Energy Commission recommends that members of the general public should not receive more than 100 millirem per year. Chronic doses may cause long-term cytotoxic and genetic effects, for example manifesting as an increased risk of a radiation-induced cancer developing later in life. Recommended limits for chronic exposure to ionizing radiation are given in Table 2.
TABLE 2Annual OccupationalOrgan or SubjectDose in milliremWhole Body  5000Lens of the Eye15,000Hands and wrists50,000Any individual organ50,000Pregnant worker500/9 monthsMinor (16-18) receiving training  100
By way of comparison, Table 3 sets forth the radiation doses from common sources.
TABLE 3SourcesDose In MilliremTelevision<1/yrGamma Rays, Jet Cross Country1Mountain Vacation - 2 week3Atomic Test Fallout5U.S. Water, Food & Air (Average)30/yrWood50/yrConcrete50/yrBrick75/yrChest X-Ray100Cosmic Radiation (Sea Level)40/yr (add 1 millirem/100 ft elev.)Natural Background San Francisco120/yrNatural Background Denver50/yrAtomic Energy Commission Limit5000/yrFor WorkersComplete Dental X-Ray5000Natural Background at Pocos de7000/yrCaldras, BrazilWhole Body Diagnostic X-Ray100,000Cancer Therapy500,000 (localized)Radiation Sickness-Nagasaki125,000 (single doses)LD50 Nagasaki & Hiroshima400,000-500,000 (single dose)
Chronic doses of greater than 5000 millirem per year (0.05 Gy per year) may result in long-term cytotoxic or genetic effects similar to those described for persons receiving acute doses. Some adverse cytotoxic or genetic effects may also occur at chronic doses of significantly less than 5000 millirem per year. For radiation protection purposes, it is assumed that any dose above zero can increase the risk of radiation-induced cancer (i.e., that there is no threshold). Epidemiologic studies have found that the estimated lifetime risk of dying from cancer is greater by about 0.04% per rem of radiation dose to the whole body.
While anti-radiation suits or other protective gear may be effective at reducing radiation exposure, such gear is expensive, unwieldy, and generally not available to public. Moreover, radioprotective gear will not protect normal tissue adjacent a tumor from stray radiation exposure during radiotherapy. What is needed, therefore, is a practical way to protect subjects who are scheduled to incur, or are at risk for incurring, exposure to ionizing radiation. In the context of therapeutic irradiation, it is desirable to enhance protection of normal cells while causing tumor cells to remain vulnerable to the detrimental effects of the radiation. Furthermore, it is desirable to provide systemic protection from anticipated or inadvertent total body irradiation, such as may occur with occupational or environmental exposures, or with certain therapeutic techniques.
Pharmaceutical radioprotectants offer a cost-efficient, effective and easily available alternative to radioprotective gear. However, previous attempts at radioprotection of normal cells with pharmaceutical compositions have not been entirely successful. For example, cytokines directed at mobilizing the peripheral blood progenitor cells confer a myeloprotective effect when given prior to radiation (Neta et al., Semin. Radiat. Oncol. 6:306-320, 1996), but do not confer systemic protection. Other chemical radioprotectors administered alone or in combination with biologic response modifiers have shown minor protective effects in mice, but application of these compounds to large mammals was less successful, and it was questioned whether chemical radioprotection was of any value (Naisin, J. R., Bacq and Alexander Award Lecture. “Chemical radioprotection: past, present, and future prospects”, Int J Radiat Biol. 73:443-50, 1998). Pharmaceutical radiation sensitizers, which are known to preferentially enhance the effects of radiation in cancerous tissues, are clearly unsuited for the general systemic protection of normal tissues from exposure to ionizing radiation.
What is needed are therapeutic agents to protect subjects who have incurred, or are at risk for incurring exposure to ionizing radiation. In the context of therapeutic irradiation, it is desirable to enhance protection of normal cells while causing tumor cells to remain vulnerable to the detrimental effects of the radiation. Furthermore, it is desirable to provide systemic protection from anticipated or inadvertent total body irradiation, such as may occur with occupational or environmental exposures, or with certain therapeutic techniques.
Protection from Toxic Side Effects of Experimental Chemotherapy
Experimental chemotherapy has been the mainstay of treatment offered to patients diagnosed with surgically unresectable advanced cancers, or cancers refractory to standard chemotherapy and radiation therapy. Of the more effective classes of drugs, curative properties are still limited. This is because of their relatively narrow therapeutic index, restricted dosage, delayed treatments and a relatively large proportion of only partial tumor reductions. This state is usually followed by recurrence, increased tumor burden, and drug resistant tumors.
Several cytoprotective agents have been proposed to enhance the therapeutic index of anticancer drugs. For methotrexate toxicity, such agents include asparaginase, leucovorum factor, thymidine, and carbipeptidase. Because of the extensive use of anthracyclines, specific and non-specific cytoprotective agents have been proposed which have varying degrees of efficacy; included are corticosteroids, desrazoxane and staurosporin. The latter is of interest in that it includes a G1/S restriction blockade in normal cells. (Chen et al., Proc AACR 39:4436A, 1998).
Cisplatin is widely used and has a small therapeutic index which has spurred investigation and search of cytoprotectants. Among the cytoprotectants for cisplatin with clinical potential are mesna, glutathione, sodium thiosulfate, and amifostine (Griggs, Leuk. Res. 22 Suppl 1:S27-33, 1998; List et al., Semin. Oncol. 23(4 Suppl 8):58-63, 1996; Taylor et al., Eur. J Cancer 33(10):1693-8, 1997). None of these or other proposed cytoprotectants such as oxonic acid for fluoropyrimidine toxicity, or prosaptide for paclitaxel PC12 cell toxicity, appears to function by a mechanism which renders normal replicating cells into a quiescent state.
What is needed are effective cytoprotective agents which are effective in protecting animals, inclusive of humans, from the cytotoxic side effects of chemotherapeutic agents.
The aryl and heteroaryl propene amide compounds of the present invention inhibit tumor cell proliferation by inducing tumor cell death without killing normal cells at therapeutically useful concentrations. The compounds of the present invention are effective against a broad range of tumor types. Without wishing to be bound by any theory, it is believed that the compounds affect the Mitogen Activated Protein Kinase (MAPK) signal transduction pathway, thereby affecting tumor cell growth and viability.